Development and characterization of monoterpene loaded microemulsions as novel scolicidal agents.

Cystic echinococcosis (CE) is one of the most important zoonotic diseases. The primary treatment is surgery and chemical sterilization of the parasitic layers by injection of a scolicidal agent. Available scolicidals possess side effects, and may cause postoperative complications. Several studies reported the scolicidal properties of monoterpene phenols and alcohols such as carvacrol, thymol, and geraniol. The present study aimed to develop, characterize, and assess monoterpene loaded microemulsions as novel green scolicidals products. For this purpose, microemulsions composing 0.37%, 0.75%, and 1.5% of monoterpenoid(s), thymol, carvacrol, and geraniol, alone or in binary or ternary mixtures were formulated. Samples were analyzed by visual inspection, polarizing optical microscope, and dynamic light scattering (DLS). The stability of the samples was evaluated up to a 3-month storage. For the scolicidal bioassay, samples at different concentrations of 200, 100, 50, 25, and 10 µg/ml were added to wells containing 104 viable protoscoleces and mortality rates were recorded at 2, 5, 10, and 20 min after exposure. Results of the present study showed that microemulsions formulated with 0.75% of pure carvacrol or the binary mixture of thymol and carvacrol at 0.375% are promising scolicidal agents.

Efficacy of niclosamide on the intra-abdominal inflammatory environment in endometriosis.

Endometriosis, a common gynecological disease, causes chronic pelvic pain and infertility in women of reproductive age. Due to the limited efficacy of current therapies, a critical need exists to develop new treatments for endometriosis. Inflammatory dysfunction, instigated by abnormal macrophage (MΦ) function, contributes to disease development and progression. However, the fundamental role of the heterogeneous population of peritoneal MΦ and their potential druggable functions is uncertain. Here we report that GATA6-expressing large peritoneal MΦ (LPM) were increased in the peritoneal cavity following lesion induction. This was associated with increased cytokine and chemokine secretion in the peritoneal fluid (PF), as well as MΦ infiltration, vascularization and innervation in endometriosis-like lesions (ELL). Niclosamide, an FDA-approved anti-helminthic drug, was effective in reducing LPM number, but not small peritoneal MΦ (SPM), in the PF. Niclosamide also inhibits aberrant inflammation in the PF, ELL, pelvic organs (uterus and vagina) and dorsal root ganglion (DRG), as well as MΦ infiltration, vascularization and innervation in the ELL. PF from ELL mice stimulated DRG outgrowth in vitro, whereas the PF from niclosamide-treated ELL mice lacked the strong stimulatory nerve growth response. These results suggest LPM induce aberrant inflammation in endometriosis promoting lesion progression and establishment of the inflammatory environment that sensitizes peripheral nociceptors in the lesions and other pelvic organs, leading to increased hyperalgesia. Our findings provide the rationale for targeting LPM and their functions with niclosamide and its efficacy in endometriosis as a new non-hormonal therapy to reduce aberrant inflammation which may ultimately diminish associated pain.

Copper nanoparticles: Biosynthesis, characterization, and protoscolicidal effects alone and combined with albendazole against hydatid cyst protoscoleces.

BACKGROUND: Surgery remains the preferred treatment option for hydatid cyst (cystic echinococcosis); however, recent studies have demonstrated that the current protoscolicidal agents used during surgery are associated with some adverse side effects such as biliary fibrosis, hepatic necrosis, and cirrhosis. The present study aims to evaluate the in vitro and ex vivo anti-parasitic effects of copper nanoparticles (CuNPs) alone and combined with albendazole on hydatid cyst protoscoleces. METHODS: CuNPs was green synthesized using C. spinosa extract. Various concentrations of CuNPs (250, 500, and 750 mg/mL) alone and combined with albendazole (ALZ, 200 mg/mL) were exposed to protoscoleces collected from the liver fertile hydatid cysts of infected sheep for 5-60 min in vitro and ex vivo. Next, the eosin exclusion test was applied to determine the viability of protoscoleces. Caspase-3 like activity of CuNPs-treated protoscoleces was then evaluated using the colorimetric protease assay Sigma Kit based on the manufacturer's instructions. RESULTS: Scanning electron microscopy (SEM) results showed that the particle size of CuNPs was 17 and 41 nm with the maximum peak at the wavelength of 414 nm. The maximum protoscolicidal activity of CuNPs was observed at the concentration of 750 mg/mL in vitro, so that 73.3 % of protoscoleces were killed after 60 min of exposure. Meanwhile, the mortality of protoscoleces was 100 % after 10 min of exposure to 750 mg/mL of CuNPs along with ALZ (200 mg/mL). Nevertheless, the findings proved that CuNPs even in combination with ALZ required a longer time to kill protoscoleces ex vivo. After 48 h of treating protoscoleces, CuNPs in a dose-dependent manner and at doses of 250, 500, and 750 mg/mL induced the caspase enzyme activation by 20.5 %, 32.3 %, and 36.1 %, respectively. CONCLUSION: The findings of the present investigation showed potent protoscolicidal effects of CuNPs, especially combined with albendazole, which entirely eliminated the parasite after 10-20 min of exposure. The results also showed that although the possible protoscolicidal mechanisms of CuNPs are not clearly understood, the inducing apoptosis through caspases is one of the main protoscolicidal mechanisms of CuNPs. However, supplementary studies, especially in animal models and clinical settings, are needed to approve these results.

Effect of Taxus baccata L. Extract on Hydatid Cyst Protoscolices In vitro.

Hydatidosis is the most important global parasitic infectious disease both in humans and animals, which can lodge at different organs of the host, such as liver, lung (even heart), and brain which may lead to death. Surgery is the main method for the treatment of hydatidosis. In surgical therapy of hydatidosis, the use of sporicidal agents is very important since these agents inactivate live protoscolices and prevent recurrence of infection. Presently, numerous scolicidal chemical agents have been administrated to inactivate the hydatid cyst contents. Recently, there has been a high tendency among researchers to evaluate and present herbal plants as alternative option due to inexpensiveness, availability, low side effects, and toxicity. This study aimed to evaluate the scolicidal effect of hydro alcoholic Taxus baccata L. extract in vitro for the first time. The scolicidal activities of the extract were tested in concentrations of 50, 100, and 150 mg/ml following 10, 30, and 60 min of incubation, and the experiments were performed in triplicate. Viability of protoscolices was confirmed by 0.1% eosin vital staining. The data were analyzed in SAS software (version 9.4). The results showed that the hydroalcoholic extract of Taxus baccata L. at the concentration of 150 mg/ml led to killing 66.6% of protoscolices at 60 min. according to the results of this investigation, it is recommended to use this plant as a scolicidal plant. The findings of the present study showed that Taxus baccata L. had potent scolicidal effects. However, further studies are required to evaluate the efficacy of Taxus baccata L. in vivo.

Effects of Allium sativum on IFN-γ and IL4 concentrations in mice with cystic echinococcosis.

The aim of the current investigation was to assess the impacts of methanolic extract of Allium sativum (MEAS) on IL-4 (a cytokine derived from Th2 cells) and IFN-É£ (a cytokine derived from Th1 cells) levels in mice infected with Echinococcus granulosus. Sixty healthy BALB/c female mice were used in this study. Each animal was intraperitoneally injected with 1500 protoscoleces. The infected animals were randomly divided into six groups: albendazole (100 mg/kg), MEAS 10 (10 mg/kg), MEAS 20 (20 mg/kg), MEAS 40 (40 mg/kg), MEAS 80 (80 mg/kg) and control group with no treatment. The studied animals received albendazole and/or MEAS through drinking water for 30 days. Serum IFN-γ concentration significantly increased in the MEAS 20 and 80 groups in comparison to the control, albendazole and MEAS 10 groups (P < 0.05). The serum IL-4 level showed no significant difference between the trial groups. The findings of this study showed that MEAS at 20 and 80 mg/kg concentrations enhanced Th1 cell response in mice with cystic echinococcosis.

Short communication: Efficacy of albendazole in Echinococcus multilocularis-infected mice depends on the functional immunity of the host.

Alveolar echinococcosis (AE) is a deadly parasitic disease that requires lifelong treatment with albendazole. Development of host immunity is pivotal with regard to the clinical outcome of AE, but its influence on conventional albendazole treatment is unknown. Using T-cell deficient athymic nude mice, we demonstrated that functional immunity is required for albendazole to be efficacious against murine AE. These results call for attention given the increasing number of immunocompromised patients with AE.

Drug repurposing applied: Activity of the anti-malarial mefloquine against Echinococcus multilocularis.

The current chemotherapeutical treatment against alveolar echinococcosis relies exclusively on benzimidazoles, which are not parasiticidal and can induce severe toxicity. There are no alternative treatment options. To identify novel drugs with activity against Echinococcus multilocularis metacestodes, researchers have studied potentially interesting drug targets (e.g. the parasite's energy metabolism), and/or adopted drug repurposing approaches by undertaking whole organism screenings. We here focus on drug screening approaches, which utilize an in vitro screening cascade that includes assessment of the drug-induced physical damage of metacestodes, the impact on metacestode viability and the viability of isolated parasite stem cells, structure-activity relationship (SAR) analysis of compound derivatives, and the mode of action. Finally, once in vitro data are indicative for a therapeutic window, the efficacy of selected compounds is assessed in experimentally infected mice. Using this screening cascade, we found that the anti-malarial mefloquine was active against E. multilocularis metacestodes in vitro and in vivo. To shed more light into the mode of action of mefloquine, SAR analysis on mefloquine analogues was performed. E. multilocularis ferritin was identified as a mefloquine-binding protein, but its precise role as a drug target remains to be elucidated. In mice that were infected either intraperitoneally with metacestodes or orally with eggs, oral treatment with mefloquine led to a significant reduction of parasite growth compared to the standard treatment with albendazole. However, mefloquine was not acting parasiticidally. Assessment of mefloquine plasma concentrations in treated mice showed that levels were reached which are close to serum concentrations that are achieved in humans during long-term malaria prophylaxis. Mefloquine might be applied in human AE patients as a salvage treatment. Future studies should focus on other repurposed anti-infective compounds (MMV665807, niclosamide, atovaquone), which showed stronger in vitro activity against E. multilocularis than mefloquine.

In vitro efficacy study of Cinnamomum zeylanicum essential oil and cinnamaldehyde against the larval stage of Echinococcus granulosus.

Hydatidosis or cystic echinococcosis is a disease caused by the larval stage of Echinococcus granulosus sensu lato. Chemotherapy can be used alone or in combination with surgery or percutaneous treatment. Benzimidazoles are the only agents used and approved for treatment, but their efficacy is extremely variable. Therefore, it is necessary to find new drugs to improve the treatment of this disease. In the last decades, the biological properties of essential oils and their components began to be investigated as alternatives in the treatment of different ailments. The aim of the present work was to evaluate the in vitro efficacy of the essential oil of Cinnamomum zeylanicum (cinnamon) and cinnamaldehyde against protoscoleces and metacestodes of E. granulosus. The essential oil and cinnamaldehyde, its major component, showed a dose and time dependent effect against protoscoleces. However, cinnamaldehyde showed a greater protoscolicidal effect than the essential oil. The maximum protoscolicidal effect was found with 50 µg/mL of cinnamaldehyde. Viability decreased by 1.7 ± 0.8% after 4 days of incubation and reached 0% at 8 days. Interestingly, there were no significant differences between the activity of cinnamaldehyde at the concentrations of 25 and 10 µg/mL and the efficacy observed with the essential oil at 200 and 50 µg/mL, respectively. Cinnamaldehyde also had a strong in vitro effect against murine cysts, while only the higher concentration of the essential oil caused ultrastructural alterations. Working with components instead of with essential oils has some advantages, particularly in relation to the reproducibility of the formulations and their effectiveness. For this reason, the results obtained in this work are promising in the search for pharmaceutical alternatives for the treatment of cystic echinococcosis.

Subcutaneous Hydatid Cyst in Laboratory Mice: Is it a Suitable Method for Evaluating Therapeutic Agents against Hydatid Cyst?

Hydatid disease is an economic and public health concern in many countries. Currently, surgery is the main treatment option for hydatid disease. In the surgical treatment of hydatidosis, the use of scolicidal agents is very important due to inactivating live protoscoleces and preventing the recurrence of infection. Therefore, it is necessary to investigate newscolicidal agents and novel medications with higher safety and efficacy. In the previous in vitro studies, the scolicidal effects of the methanolic extracts and aromatic water of Zataria multiflora (Z. multiflora) have been demonstrated. Consequently, in this study, the impact of the nanoemulsion of Z. multiflora essential oil on subcutaneous hydatid cysts was compared with albendazole (ABZ). Fifty laboratory male mice were inoculated with 300 viable protoscoleces subcutaneously on the two sides of the abdomen. Following five months of infection, the remaining infected mice (n=42) were allocated into two treatment and one control (without treatment) groups containing fourteen animals each. Group A received ABZ at the dose of 50 mg/kg for 60 days, group B received the nanoemulsions of Z. multiflora at the dose of 50 mg/kg in drinking water for 60 days, and group C was considered as the control group. All the infected mice were euthanized and necropsied two months post-intervention. Afterwards, the cysts were cautiously collected and their number, size, and weight were compared between the mice of different groups. The mean number of hydatid cysts indicated that the nanoemulsion of Z. multiflora essence had a relative superiority to ABZ. On the other hand, the therapeutic effect of ABZ was higher than the nanoemulsion of Z. multiflora essential oil in terms of the mean weight and mean size of hydatid cysts. However, no significant difference was observed between the groups (P&gt;0.5). Overall, the number, weight, and size of cysts were not significantly different between the groups in this investigation. The lack of satisfactory therapeutic results in this study might be due to the location of hydatid cysts in the subcutaneous space.

Histopathologic evaluation of experimental murine neurocysticercosis after treatment with albendazole/nitazoxanide combination.

Neurocysticercosis (NCC) is the most common helminthic brain infection related to epilepsy. Only albendazole (ABZ) and praziquantel are used in its treatment. The development of new therapeutics has been encouraged. Taenia crassiceps cysticerci intracranial infection is the experimental model used in NCC studies. This study evaluated the histopathology of the brains of BALB/c mice experimentally infected with T. crassiceps cysticerci after the treatment with the ABZ/nitazoxanide (NTZ) combination. Thirty days after the inoculation the mice received an oral single dose of the ABZ/NTZ combination (40 mg kg-1 each). The control groups were treated with: NaCl 0.9%; ABZ or NTZ. The histopathologic evaluation of the brains was performed 24 h after treatment. The ABZ treatment induced discrete mononuclear inflammatory infiltration, meningitis, gliosis, hyperaemia and hippocampus compression; moderate ependimitis and oedema. The NTZ treatment induced accentuated inflammatory infiltration, foamy macrophages, ependimitis, choroiditis, gliosis and hyperaemia and moderate oedema. The ABZ/NTZ combination treatment induced a significant decrease in the polymorphonuclear inflammatory infiltration, ependimitis, choroiditis, gliosis, hyperaemia and ventriculomegaly in comparison with the other groups. The cysticerci showed destruction of the tegument not observed in other groups. The ABZ/NTZ combination is efficient as the parasite showed signs of destruction and lower damage to the host's tissue.

Scolicidal Effects of Nanoparticles Against Hydatid Cyst Protoscolices in vitro.

BACKGROUND: Echinococcus granulosus is causative agent of cystic echinococcosis (CE), which has a cosmopolitan distribution. The current methods for the treatment of human CE include surgery. Therefore, the development of new scolicidal agents with low side effects and more efficacies is an urgent need. PURPOSE: The present study aimed to compare the scolicidal efficacies of silver, iron, copper, silica and zinc oxide nanoparticles (NPs) against hydatid cyst protoscolices in vitro. METHODS: Hydatid cysts of sheep liver and lung were collected. The cyst fluid containing protoscolices was aspirated aseptically. The scolicidal activities of the silver, iron, copper, silica and zinc nanoparticles (Ag-NP, Fe-NP, Cu-NP, Si-NP and Zn-NP) were tested at different concentrations of 0.25, 0.5 and 1 mg/mL following 10, 30 and 60 min of incubation in triplicate. Viability of protoscolices was confirmed by 0.1% eosin staining. RESULTS: Results showed that Ag-NPs at all concentrations tested had the highest scolicidal effect. Ag-NPs at 1 mg/mL concentration after 60 min of exposure time showed 80% mortality rate. Si-NPs had the high scolicidal activity at 1 mg/mL concentration (52.33%), Cu-NPs at 0.5 mg/mL concentration (41%), Fe-NPs at 1mg/mL concentration (28%) and Zn-NPs at concentration of 1mg/mL after 60 mins (15.67%). CONCLUSION: The findings of the present study showed that Ag-NPs, Fe-NPs, Cu-NPs, Si-NPs and Zn-NPs had potent scolicidal effects and that Ag-NPs are recommended as effective scolicidal agents. However, further in vivo studies are required to evaluate the efficacy of these nanoparticles.

In vitro effects of lonidamine and 6-aminonicotinamide against Echinococcus granulosussensu stricto and Echinococcus multilocularis.

Echinococcosis is a zoonotic disease caused by cestode species of the genus Echinococcus, which demonstrates considerable medical and veterinary concerns. The development of novel drugs for echinococcosis treatment is urgently needed. In this study, we demonstrated that lonidamine (LND) and 6-aminonicotinamide (6-AN) exhibited considerable in vitro effects against both larval- and adult-stage of E. granulosussensu stricto (s. s.) and E. multilocularis. The combination of LND and 6-AN exhibited a significantly higher activity than the single drug treatment. These results highlight the therapeutic potential of LND, 6-AN and the combination of LND and 6-AN for the treatment of echinococcosis.

Niclosamide suppresses macrophage-induced inflammation in endometriosis†.

Endometriosis is a common gynecological disease, which causes chronic pelvic pain and infertility in women of reproductive age. Due to limited efficacy of current treatment options, a critical need exists to develop new and effective treatments for endometriosis. Niclosamide is an efficacious and FDA-approved drug for the treatment of helminthosis in humans that has been used for decades. We have reported that niclosamide reduces growth and progression of endometriosis-like lesions via targeting STAT3 and NFĸB signaling in a mouse model of endometriosis. To examine the effects of niclosamide on macrophage-induced inflammation in endometriosis, a total of 29 stage III-IV endometrioma samples were used to isolate human endometriotic stromal cells (hESCs). M1 or M2 macrophages were isolated and differentiated from fresh human peripheral blood samples. Then, hESCs were cultured in conditioned media (CM) from macrophages with/without niclosamide. Niclosamide dose dependently reduced cell viability and the activity of STAT3 and NFκB signaling in hESCs. While macrophage CM stimulated cell viability in hESCs, niclosamide inhibited this stimulation. Macrophage CM stimulated the secretion of proinflammatory cytokines and chemokines from hESCs. Most of these secreted factors were inhibited by niclosamide. These results indicate that niclosamide is able to reduce macrophage-induced cell viability and cytokine/chemokine secretion in hESCs by inhibiting inflammatory mechanisms via STAT3 and/or NFκB signaling.

In vitro metabolomic footprint of the Echinococcus multilocularis metacestode.

Alveolar echinococcosis (AE) is a zoonotic disease that is deadly if left untreated. AE is caused by the larval metacestode stage of the cestode Echinococcus multilocularis. Better knowledge on the host-parasite interface could yield novel targets for improvement of the treatment against AE. We analyzed culture media incubated with in vitro grown E. multilocularis metacestodes by 1H nuclear magnetic resonance spectroscopy to identify the unknown metabolic footprint of the parasite. Moreover, we quantitatively analyzed all amino acids, acetate, glucose, lactate, and succinate in time-course experiments using liquid chromatography and enzymatic assays. The E. multilocularis metacestodes consumed glucose and, surprisingly, threonine and produced succinate, acetate, and alanine as major fermentation products. The metabolic composition of vesicle fluid (VF) from in vitro grown E. multilocularis metacestodes was different from parasite-incubated culture medium with respect to the abundance, but not the spectrum, of metabolites, and some metabolites, in particular amino acids, accumulated in the VF. Overall, this study presents the first characterization of the in vitro metabolic footprint of E. multilocularis metacestodes and VF composition, and it provides the basis for analyses of potentially targetable pathways for future drug development.

In vitro and in vivo effects of 3-bromopyruvate against Echinococcus metacestodes.

While searching for novel anti-echinococcosis drugs, we have been focusing on glycolysis which is relied on by Echinococcus for energy production and intermediates for other metabolic processes. The aim of this study was to investigate the potential therapeutic implication of glycolytic inhibitors on Echinococcus. Our results demonstrate that at an initial concentration of 40 µM, all inhibitors of glycolysis used in the current experiment [3-bromopyruvate (3-BrPA), ornidazole, clorsulon (CLS), sodium oxamate and 2,6-dihydroxynaphthalene (NA-P2)] show considerable in vitro effects against Echinococcus granulosus protoscoleces and Echinococcus multilocularis metacestodes. Among them, 3-BrPA exhibited the highest activity which was similar to that of nitazoxanide (NTZ) and more efficacious than albendazole (ABZ). The activity of 3-BrPA was dose dependent and resulted in severe ultrastructural destructions, as visualized by electron microscopy. An additional in vivo study in mice infected with E. multilocularis metacestodes indicates a reduction in parasite weight after the twice-weekly treatment of 25 mg/kg 3-BrPA for 6 weeks, compared to that of the untreated control. In particular, in contrast to ABZ, the administration of 25 mg/kg 3-BrPA did not cause toxicity to the liver and kidney in mice. Similarly, at the effective dose against Echinococcus larvae, 3-BrPA showed no significant toxicity to human hepatocytes. Taken together, the results suggest that interfering with the glycolysis of the parasite may be a novel chemotherapeutical option and 3-BrPA, which exhibited a remarkable activity against Echinococcus, may be a promising potential drug against cystic echinococcosis (CE) and alveolar echinococcosis (AE).

In vitro efficacy of ampelopsin against Echinococcus granulosus and Echinococcus multilocularis.

The metacestode stage of Echinococcus granulosus and Echinococcus multilocularis cause cystic echinococcosis and alveolar echinococcosis, respectively, which result in severe medical and veterinary problems. In this study, as an exploration of novel treatment agents against echinococcosis, we demonstrated that ampelopsin (AMP), which is extracted from Ampelopsis grossedentata and has been clinically used for treatments of various types of diseases including cancers for a long time, exhibited profound in vitro effect against E. granulosus protoscoleces and E. multilocularis metacestodes. Furthermore, in vitro cytotoxicity assay also demonstrated that AMP at the effective dose against E. granulosus protoscoleces and E. multilocularis metacestodes did not show significant toxicity to human hepatocytes. These results suggest that AMP has the potential as an alternative agent against echinococcosis.

Scolicidal activity of Mesobuthus eupeus venom against the protoscolices of Echinococcus granulosus.

Hydatidosis is an important zoonosis caused by a parasitic tapeworm, namely Echinococcus granulosus. This infection is distributed worldwide and affects the health as well as economic loss in both humans and animals. In most cases, the disease needs chemotherapy with or without surgery. Conventional drugs have some major problems, including drug complications, harmful side effects, and also progressive resistance. According to the importance of biological productions as alternative medicine, a large number of studies confirmed that whole venom and many peptide ingredients of the scorpion venom have various different medical benefits, including antimicrobial properties, due to the mechanism of blocking gated ion channel. In this study, the venom peptides of Mesobuthus eupeus scorpionwere purified using gel filtration chromatography and subsequently ion exchange chromatography, followed by the determination of the molecular weights of the proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) procedure. After collecting the hydatid cysts fluids from the liver of infected sheep, protoscolices were derived, washed, and encountered to the whole venom as well as eight different fractions of toxin 30, 60, 120, and 240 min after the exposure. In the next step, the viability of protoscolices was determined by eosin staining. The obtained results revealed that a venom fraction under 10 kDa killed all protoscolices after 30 min. Moreover, it was found that the scolicidal activity of fractions increases according to the time of exposure. As a result, it can be concluded that M. epeus venom peptides under its LD50 (1/2 LD50) can properly and quickly destroy the protoscolices of hydatid cysts at the level of applied concentrations and such components are good alternatives to treat hydatidosis.

Scolicidal Effects of Chitosan-Curcumin Nanoparticles on the Hydatid Cyst Protoscolices.

PURPOSE: In the current era, cystic echinococcosis (CE), as larval stage of Echinococcus granulosus, is considered as a threat to human health. Scolicidal agents used in the surgery of cysts have different side effects. Therefore, the present study aimed to assess the effects of chitosan nanoparticles containing curcumin (Ch-Cu NPs) on the protoscolices of the hydatid cyst in vitro. METHODS: Ch-Cu NPs were synthesized using a simple co-precipitation method and their structural and morphological properties were characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM), zeta analyzer, and Fourier transform infrared (FT-IR) spectroscopy. Then, the effects of different concentrations of Ch-Cu NPs (0.25, 0.05, 1, 2, and 4 mg/mL) on the fatality rate, and the length and width of protoscolices in different times (5, 10, 20, 30, and 60 min) were investigated. In addition, the SEM technique was used to evaluate the structure of the protoscolices after treatment. RESULTS: Based on the results, the presence of curcumin on the chitosan nanoparticles was confirmed by FT-IR analysis. Further, XRD analysis approved the crystal structure of chitosan NPs. Furthermore, the highest fatality rate was 68% in 4 mg/mL concentration of Ch-Cu NPs. The length and width of protoscolices decreased based on the high concentrations of Ch-Cu NPs, compared to the control group. CONCLUSION: Finally, Ch-Cu NPs expressed good scolicidal activities, which made them suitable to be considered as an anti-protoscolex agent.

In vitro evaluation of the protoscolicidal effect of Eucalyptus globulus essential oil on protoscolices of hydatid cyst compared with hypertonic saline, povidone iodine and silver nitrate.

OBJECTIVE: There are various protoscolicidal agents for inactivation of protoscoleces of hydatid cysts before and during surgical operation. The present study was aimed to evaluate the protoscolicidal effect of two concentrations of Eucalyptus globulus on protoscoleces of Echinococcus granulosus sensu lato under in vitro condition and to compare its efficacy with hypertonic saline, povidone iodine and silver nitrate. METHODS: Live protoscoleces obtained from the liver of naturally infected sheep were exposed to 0.5% and 1% of Eucalyptus globulus essential oil, 5% hypertonic saline, 10% povidone iodine and 0.5% silver nitrate for 1 and 3minutes. Phosphate buffered saline was used as a negative control. One percent eosin staining method was used to test the viability of protoscoleces in different groups. RESULTS: While the mean percentage of dead protoscoleces was 6.08% in the control group, the scolicidal power of 5% hypertonic saline was only 6.54% and 6.60% after 1 and 3min respectively. 0.5% E. globulus EO demonstrated 97.38% and 100% scolicidal activity after 1 and 3min respectively. The mean protoscolicidal power of 1% E. globulus EO, 10% povidone iodine and 0.5% silver nitrate was 100% after one minute. CONCLUSIONS: According to the results of this study, E. globulus EO demonstrated high scolicidal power in a short period of time. Hence, this herbal product could be considered as a potent natural scolicidal agent that could be used before and during surgery of hydatid disease.

Sodium arsenite augments sensitivity of Echinococcus granulosus protoscoleces to albendazole.

This study aimed to observe the effects of sodium arsenite (NaAsO2) on apoptosis of Echinococcus granulosus protoscoleces induced by albendazole (ABZ), and to explore the potential mechanism of NaAsO2. According to the following final concentrations, the experimental groups were divided into 10 µM NaAsO2, 20 µM NaAsO2, 80 µM ABZ, 10 µM NaAsO2+80 µM ABZ, and 20 µM NaAsO2+80 µM ABZ. Viability was detected with 0.1% eosin staining. The ultrastructural alterations were visualized by scanning electron microscopy. Caspase-3 activity was assessed with colorimetric assay. Meanwhile, ELISA or WST were applied to detect the activities of antioxidases in NaAsO2 treatment groups. The maximum protoscolicidal effect was seen with the combination 20 µM NaAsO2+80 µM ABZ. The ultrastructural damage detected after NaAsO2+ABZ incubation were greater than those caused by ABZ alone and its primary damage site was the tegument of the parasite. The caspase-3 activity was clearly higher in protoscoleces treated with the combination of NaAsO2+ABZ than when drugs were used separately. The activities of NQO-1, HO-1, GST, and SOD were significantly lower in protoscoleces incubated with NaAsO2 than the untreated controls (P < 0.05). According to our results, ABZ could induce protoscoleces apoptosis, and NaAsO2 could significantly augment sensitivity of protoscoleces to ABZ.